Mohandas A R, Dekaban G A, Dales S
Department of Microbiology and Immunology, University of Western Ontario, London, Canada.
Virology. 1994 Apr;200(1):207-19. doi: 10.1006/viro.1994.1179.
Polypeptide Ag35, a major early component of the vaccinia surface, is integrated into the formative viral lipoprotein tegument. To ascertain whether positioning of Ag35 is due to its general affinity for newly assembled viral membranes we created a recombinant A12 vector to express the vaccinia protein. The baculovirus system was chosen because intranuclear virions of this agent are likewise enclosed inside newly formed envelopes. Comparable infections of two insect cell lines established that more abundant synthesis occurred in High Five (H5) than in SF9 cells. We, therefore, used H5 cells for most experiments reported here. Combined analyses by PAGE, Western blotting, and immunocytology, using light and electron microscopy, revealed a dissemination of Ag35 throughout the cell. Higher concentrations were evident at the cell surface, nuclear perimeter, and within intranuclear virogenic stroma. The association with the virogenic stroma was of specific interest with respect to vaccinia development because it showed a similarity in the targeting of Ag35 toward intranuclear DNA-protein foci of baculovirus which are analogous to the vaccinia-specified cytoplasmic "factories." A further remarkable analogy concerns association of Ag35 with intranuclear baculovirus envelopes, revealing a propensity of Ag35 for nascent viral lipoprotein membranes.
多肽Ag35是痘苗病毒表面的一种主要早期成分，它整合到正在形成的病毒脂蛋白内膜中。为了确定Ag35的定位是否归因于其对新组装的病毒膜的一般亲和力，我们构建了一种重组A12载体来表达痘苗病毒蛋白。选择杆状病毒系统是因为该病原体的核内病毒粒子同样被包裹在新形成的包膜内。对两种昆虫细胞系进行的类似感染实验表明，在High Five（H5）细胞中合成的量比在SF9细胞中更多。因此，我们在本文报道的大多数实验中使用了H5细胞。通过PAGE、蛋白质印迹和免疫细胞化学方法，并结合光学显微镜和电子显微镜进行的综合分析显示，Ag35在整个细胞中都有分布。在细胞表面、核周以及核内病毒发生基质中可以明显看到较高的浓度。就痘苗病毒的发育而言，Ag35与病毒发生基质的关联特别令人感兴趣，因为这表明Ag35靶向杆状病毒的核内DNA - 蛋白质聚集区，这类似于痘苗病毒指定的细胞质“工厂”。另一个显著的相似之处是Ag35与核内杆状病毒包膜的关联，这揭示了Ag35对新生病毒脂蛋白膜的倾向。
